We have studied the phototoxicity of gamma-cyclodextrin bicapped pristine C-60 (gamma-CyD)(2)/C-60 and its water-soluble derivative C-60(OH)(24) (fullerol) toward human keratinocytes. Our results demonstrated that irradiation of (gamma-CyD)(2)/C-60 or fullerol in D2O generated singlet oxygen with quantum yields of 0.76 and 0.08, respectively. Irradiation (>400 nm) of fullerol generated superoxide as detected by the EPR spin trapping technique;superoxide generation was enhanced by addition of the electron donor nicotinamide adenine dinucleotide (reduced) (NADH). During the irradiation of (gamma-CyD)(2)/C-60, superoxide was generated only in the presence of NADH. Cell viability measurements demonstrated that (y-CyD)(2)/C-60 was about 60 times more phototoxic to human keratinocytes than fullerol. UVA irradiation of human keratinocytes in the presence of (gamma-CyD)(2)/C-60 resulted in a significant rise in intracellular protein-derived peroxides, suggesting a type II mechanism for phototoxicity. UVA irradiation of human keratinocytes in the presence of fullerol produced diffuse intracellular fluorescence when the hydrogen peroxide probe Peroxyfluor-1 was present, suggesting a type I mechanism. Our results show that the phototoxicity induced by (gamma-CyD)(2)/C-60 is mainly mediated by singlet oxygen with a minor contribution from superoxide, while fullerol phototoxicity is mainly due to superoxide. We also examined the effect of (gamma-CyD)(2)/C-60 on human lens epithelial cells exposed to light, and found that this water soluble fullerene damages lens cells primarily through a singlet oxygen mechanism. We determined the cytotoxicity and visible light induced phototoxicity of fullerol in vitro with human retinal pigment epithelial (hRPE) cells, finding that fullerol binds to hRPE cells and causes their apoptosis and necrosis, with the cytotoxic effects increasing with fullerol concentration. Fullerol decreased mitochondrial activity and led to cell membrane damage in hRPE cells in the dark at concentrations of 20-50 M, as shown by MTS and LDH assays, respectively. The cytotoxicity increased when cells preincubated with fullerol were exposed to visible light. Increased TBARS production induced by light was correlated with fullerol concentration in hRPE cells. Thus fullerol is both cytotoxic in the dark and phototoxic to hRPE cells at low concentrations and weak visible light, factors that might limit their use as drug delivery systems. The mechanism of this damage is, at least in part, through singlet oxygen production resulting in lipid peroxidation, an important risk factor in the induction of retinal and choroidal neovascularization and diabetic retinopathy. We also investigated the strongly red-shifted singlet oxygen phosphorescence spectra in an aqueous preparation of C-60 buckminsterfullerene. The red shift of about 10 nm was associated with aqueous dispersions of C-60 nanoaggregates (C-60)n that can photosensitize singlet oxygen in their polarizable cores. In contrast to singlet oxygen produced by the water-soluble C-60-(c-cyclodextrin)2 complex, singlet oxygen trapped inside (C-60)n was short-lived (on the order of 2 -3 microseconds) and insensitive to solvent and singlet oxygen quenchers. It did not induce photocytotoxicity. To our knowledge, the singlet oxygen spectrum from (C-60)n is the most red-shifted singlet oxygen spectrum recorded to date, and it may be useful for probing the inner polarizability of carbon (nano) aggregates.